气肿疽梭菌套式PCR检测方法的建立  

Establishment of a nested PCR assay for detection of Clostridium chauvoei

作　　者：姜丹丹[1] 金鑫[1] 陈莹[1] 车达[1] 费景春 

机构地区：[1]延边大学农学院动物医学系,吉林延吉133000 [2]吉林松原职业技术学院,吉林松原138000

出　　处：《中国兽医科学》2010年第11期1171-1174,共4页Chinese Veterinary Science

基　　金：吉林省延边州科技计划项目(2008YB501A02)

摘　　要：根据GenBank中的气肿疽梭菌16SrRNA基因序列(登录号EU106373),设计合成了2对特异性引物,建立了适合气肿疽梭菌快速检测的套式PCR方法。对该方法的最佳反应条件进行了筛选,并采用该方法对气肿疽梭菌进行了检测。结果显示,气肿疽梭菌能扩增到341bp的条带,而魏氏梭菌、巴氏杆菌和腐败梭菌的扩增结果均为阴性。该方法第1次扩增的敏感性是56.41pg,第2次扩增的敏感性是56.41fg,第2次比第1次扩增的敏感性高103倍。表明,建立的套式PCR方法具有较高的敏感性和特异性,可用于气肿疽梭菌的检测,为气肿疽的临床检测、流行病学调查和进出口检疫提供了新的技术手段。According to the sequence of 16S rRNA gene of Clostridium chauvoei available in GenBank（Accession No.EU106373） ,two pairs of specific primers were designed and synthesized,then a nested PCR assay for rapid detection of C.chauvoei was established.Reaction conditions of the nested PCR were optimized.A specific 341bp fragment was amplified from DNA templates of C.chauvoei,but no band was amplified with templates extracted from Clostridium welchii,Pasteurella multocida or Clostridium septicum.Sensitivity of the 1st and 2nd amplifications by the nested PCR assay was 56.41pg and 56.41fg,respectively.The sensitivity of the 2nd amplifications was increased 10 3times.These results showed that this detection method had higher sensitivity and specificity,and could be used for the detection of C.chauvoei,providing a new technology for the detection in clinical cases,epidemiological investigation and import-export quarantine of emphysematous gangrene.

关 键 词：气肿疽梭菌 16S RRNA基因 套式聚合酶链反应 检测 

分 类 号：S852.616.3[农业科学—基础兽医学]